The long-term goal of this program is to identify and develop novel inhibitors of human sphingosine kinase (SK) that are effective as therapeutic agents. Because of its critical role in sphingolipid metabolism and angiogenesis, we have focused on SK as an innovative molecular target for the development of new drugs for the treatment of diabetic retinopathy. Sphingolipids are being increasingly recognized as key mediators of apoptosis, stress responses, cell differentiation and proliferation, and are known to interact with key players in the angiogenic cascade of central importance in diabetic retinopathy. Specifically, sphingosine-1-phosphate (S1P) produced by SK mediates the effects of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and protein kinase C (PKC), all having critical roles in the manifestations of diabetic retinopathy. Therefore, SK is a key molecular target for the development of new therapeutic agents for this disease. In spite of accumulating evidence for a pivotal role of SK in regulating proliferation and angiogenesis, pharmacological inhibition of SK is an untested means of treating an angiogenic-based disease such as diabetic retinopathy. This is largely due to the heretofore lack of pharmacologically useful SK inhibitors. To overcome this problem, we have recently identified novel inhibitors of human SK. We hypothesize that these SK inhibitors will be useful to block the deleterious angiogenic effects of VEGF and bFGF, and thereby ameliorate diabetic retinopathy. To provide proof-of-principle evaluations of the utility of these compounds, the following Specific Aims will be addressed in Phase I of this project: 1). To synthesize sufficient amounts of three SK inhibitors for in vitro and in vivo studies. 2). To determine the in vitro effects of these SK inhibitors on the VEGF- and bFGF-mediated signaling cascades that contribute to angiogenesis. 3). To evaluate the in vivo toxicities and therapeutic efficacies of SK inhibitors in two rodent models of diabetic retinopathy. Because of our previous work that led to the identification of these compounds and the development of methods for their synthesis, as well as the demonstrated expertise of our Consultant with diabetic retinopathy models, we are currently in a unique position to undertake the proposed studies.